The aim of this study was to analyse the influence of the microtopography and hydrophilicity of titanium (Ti) substrates on initial oral biofilm formation.

Materials and methods

Nine bacterial species belonging to the normal oral microbiota, including: Aggregatibacter actinomycetemcomitans, Actinomyces israelii, Campylobacter rectus, Eikenella corrodens, Fusobacterium nucleatum, Parvimonas micra, Porphyromonas gingivalis, Prevotella intermedia, and Streptococcus sanguinis were tested on Ti surfaces: pretreatment (PT [Ra<0.2 μm]), acid-etched (A [Ra<0.8 μm]), A modified to be hydrophilic (modA), sand-blasted/acid-etched (SLA [Ra = 4 μm]), and hydrophilic SLA (modSLA). Disks were incubated for 24 h in anaerobic conditions using a normal culture medium (CM) or human saliva (HS). The total counts of bacteria and the proportion of each bacterial species were analysed by checkerboard DNA–DNA hybridization. Results: Higher counts of bacteria were observed on all surfaces incubated with CM compared with the samples incubated with HS. PT, SLA, and modSLA exhibited higher numbers of attached bacteria in CM, whereas SLA and modSLA had a significant increase in bacterial adhesion in HS. The proportion of the species in the initial biofilms was also influenced by the surface properties and the media used: SLA and modSLA increased the proportion of species like A. actinomycetemcomitans and S. sanguinis in both media, while the adhesion of A. israelii and P. gingivalis on the same surfaces was affected in the presence of saliva.

Conclusions

The initial biofilm formation and composition were affected by the microtopography and hydrophilicity of the surface and by the media used.

Cellular and molecular events in osseointegration at the dental implant surface remain largely unknown. We hypothesized that bone marrow stromal cells (BMSCs) participate in this process, and that osterix (Osx) promotes implant osseointegration. To prove this hypothesis, we tracked double-labeled BMSCs in implantation sites created in nude mice transplanted with these cells. We also inserted implants into the femurs of our established transgenic mice after local administration of viruses encoding Osx, to determine the osteogenic effects of Osx. Immunohistochemical results demonstrated that BMSCs can recruit from peripheral circulation and participate in wound healing and osseointegration after implantation. Microcomputed tomography (microCT) analysis revealed an increased bone density at the bone-to-implant interface in the Osx group, and histomorphometric analysis indicated an elevated level of bone-to-implant contact in the Osx group. We concluded that exogenous BMSCs participate in the osseointegration after implantation, and that Osx overexpression accelerates osseointegration.

Osteoblasts grown on microstructured Ti surfaces enhance osteointegration by producing local factors that regulate bone formation as well as bone remodeling, including the RANK ligand decoy receptor osteoprotegerin (OPG). The objective of this study was to explore the mechanism by which surface microstructure and surface energy mediate their stimulatory effects on OPG expression. Titanium disks were manufactured to present different surface morphologies: a smooth pretreatment surface (PT, Ra<0.2μm), microstructured sandblasted/acid etched surface (SLA, Ra=3-4μm), and a microstructured Ti plasma-sprayed surface (TPS, Ra=4μm). Human osteoblast-like MG63 cells were cultured on these substrates and the regulation of OPG production by TGF-β1, PKC, and α2β1 integrin signaling determined. Osteoblasts produced increased amounts of OPG as well as active and latent TGF-β1 and had increased PKC activity when grown on SLA and TPS. Exogenous TGF-β1 increased OPG production in a dose-dependent manner on all surfaces, and this was prevented by adding blocking antibody to the TGF-β type II receptor or by reducing TGF-β1 binding to the receptor by adding exogenous soluble type II receptor. The PKC inhibitor chelerythrine inhibited the production of OPG in a dose-dependent manner, but only in cultures on SLA and TPS. shRNA knockdown of α2 or a double knockdown of α2β1 also reduced OPG, as well as production of TGF-β1. These results indicate that substrate dependent OPG production is regulated by TGF-β1, PKC, and α2β1 and suggest a mechanism by which α2β1-signaling increases PKC, resulting in TGF-β1 production and TGF-β1 then acts on its receptor to increase transcription of OPG.

Cellular and molecular events in osseointegration at the dental implant surface remain largely unknown. We hypothesized that bone marrow stromal cells (BMSCs) participate in this process, and that osterix (Osx) promotes implant osseointegration. To prove this hypothesis, we tracked double-labeled BMSCs in implantation sites created in nude mice transplanted with these cells. We also inserted implants into the femurs of our established transgenic mice after local administration of viruses encoding Osx, to determine the osteogenic effects of Osx. Immunohistochemical results demonstrated that BMSCs can recruit from peripheral circulation and participate in wound healing and osseointegration after implantation. Microcomputed tomography (microCT) analysis revealed an increased bone density at the bone-to-implant interface in the Osx group, and histomorphometric analysis indicated an elevated level of bone-to-implant contact in the Osx group. We concluded that exogenous BMSCs participate in the osseointegration after implantation, and that Osx overexpression accelerates osseointegration.

Surface roughness and surface free energy are two important factors that regulate cell responses to biomaterials. Previous studies established that titanium substrates with micron-scale and submicron scale topographies promote osteoblast differentiation and osteogenic local factor production and that there is a synergistic response to microrough Ti surfaces that have retained their high surface energy via processing that limits hydrocarbon contamination. This study tested the hypothesis that the synergistic response of osteoblasts to these modified surfaces depends on both surface microstructure and surface energy.

Methods

Ti disks were manufactured to present three different surface structures: smooth pretreatment surfaces (PT) with Ra of 0.2 µm; acid-etched surfaces (A) with a submicron roughness Ra of 0.83 µm; and sandblasted/acid-etched surfaces (SLA) with Ra of 3–4 µm. Modified acid-etched (modA) and modified sandblasted/acid-etched (modSLA) titanium substrates, which have low contamination and present a hydroxylated/hydrated surface layer to retain high surface energy, were compared with regular low surface energy A and SLA surfaces. Human osteoblast-like MG63 cells were cultured on these substrates and their responses, including cell shape, growth, differentiation (alkaline phosphatase, osteocalcin), and local factor production (TGF-β1, PGE2, osteoprotegerin [OPG]) were analyzed (N=6 per variable). Data were normalized to cell number.

Results

There were no significant differences between smooth PT and A surfaces except for a small increase in OPG. Compared to A surfaces, MG63 cells produced 30% more osteocalcin on modA, and 70% more on SLA. However, growth on modSLA increased osteocalcin by more than 250%, which exceeded the sum of independent effects of surface energy and topography. Similar effects were noted when levels of latent TGF-β1, PGE2 and OPG were measured in the conditioned media.

Conclusions

The results demonstrate a synergistic effect between high surface energy and topography of Ti substrates and show that both micron scale and submicron scale structural features are necessary.

Title II of the Ryan White Comprehensive AIDS Resources Emergency (CARE) Act of 1990 provides formula-based grants to States to help them improve the quality, availability, and organization of health care and support services for people with human immunodeficiency virus (HIV) infection. This article reviews State expenditures during the first year of CARE Act funding (April 1991-March 1992) within the context of Title II guidelines and the federally funded grant programs that preceded and helped shape Title II. The authors also discuss future challenges that require development of resources, the assessment of program impact, and the evaluation of the quality and appropriateness of HIV-related services. Ninety-one percent of the $77.5 million awarded to States during fiscal year 1991 went for the provision of medical and support services through HIV care consortia, drug reimbursement programs, home and community-based care programs, and health insurance initiatives. The remaining monies were used for planning, evaluation, and program administration. Forty States allocated $38.9 million for the establishment of HIV care consortia to assess service needs and to develop comprehensive continuums of health and support services in the areas most affected by HIV disease. Fifty States allocated an additional $28.3 million for the continuation or expansion of FDA-approved drug therapies for low-income people with HIV infection. Twenty-five States allocated $2.2 million for the provision of home- and community-based health services, and 16 States allocated $1.3 million for programs that help low-income people with HIV infection to purchase or maintain health insurance coverage.